Spleen tyrosine kinase (Syk) was originally cloned from porcine spleen (Taniguchi et al., 1991, J. Biol. Chem. 266:15790-15796). It was initially purified as a 40 kD protein-tyrosine kinase (PTK), but it was later found to be a major proteolytic product of a larger 72 kD mature protein (Taniguchi et al., 1991, J. Biol. Chem. 266:15790-15796; Law et al., 1994, J. Biol. Chem. 269:12310-12319; Zioncheck et al., 1986, J. Biol. Chem. 261:15637-15643; Zioncheck et al., 1988, J. Biol. Chem. 263:19195-19202) The human Syk locus has been mapped to chromosome 9 at band q22 (Law et al., 1994, J. Biol. Chem. 269:12310-12319).
Syk was long thought to be a hematopoietic cell-specific signaling molecule (Yan et al., 1997, J. Immunol. 158:1902-1910; Qin et al., 1997, Biochem. Biophys. Res. Commun. 236:697-701; Palmieri et al., 1999, J. Immunol. 162:7181-7188; Ohta et al., 1992, Biochem. Biophys. Res. Commun 185:1128-1132; Schieven et al., 1993, J. Biol. Chem. 268:16688-16692; Chan et al., 1994, J. Immunol. 152:4758-4766; Yousefi et al., 1996, J. Exp. Med. 183:1407-1414; Benhamou et al., 1993, J. Biol. Chem. 268:23318-23324; Musch et al., 1999, J. Biol. Chem. 274:7923-7928; Harrison et al., 1994, J. Biol. Chem. 269:955-959; Zhang et al., 1996, J. Exp. Med. 184:71-79). Recent studies have demonstrated that Syk is expressed by many non-hematopoietic cells including normal and tumorigenic mammary epithelial cells (Coopman et al., 2000, Nature 406:742-747), airway epithelial cells, vascular endothelial cells, melanocytes, neuron-like cells (Ulanova et al., 2005, Am. J. Physiol. Lung Cell. Mol. Physiol. 288:L497-L507; Inatome et al., 2001, Biochem. Biophys. Res. Commun. 286:195-199; Tsujimura et al., 2001, Fed. Eur. Biochem. Soc. Lett. 489:129-133; Hoeller et al., 2005, J. Invest. Dermatol. 124:1293-1299), the mouse embryonic fibroblast cell line 3T3-L1 (where it plays a role in the differentiation of 3T3-L1 to adipocytes and adipogenesis (Wang et al., 1999, J. Biol. Chem. 274:32159-32166), and human nasal fibroblasts (Yamada et al., 2001, J. Immunol. 166:538-543).
Some data suggest that Syk could be a potential marker for tumor formation and progression. A comparison of Syk mRNA levels between primary breast cancer tissue and adjacent non-cancerous breast tissue using real-time quantitative PCR demonstrated that patients with reduced Syk expression have increased risk for distant metastasis (Toyama et al., 2003, Cancer Lett. 189:97-102). Wang et al. (2004, World J. Gastroenterol. 10:1815-1818) demonstrated that Syk expression was lower in gastric cancer patients with lymph node metastasis than in patients without lymph node metastasis. Goodman et al. (2001, Oncogene 20:3969-3978) demonstrated that reduced Syk expression and activity is observed in the leukemic cells from acute lymphoblastic leukemia (ALL) patients. The Syk mRNA sequence in pro-B leukemia cells of ALL patients exhibited deletions or insertions that result in abnormal Syk proteins with a missing or truncated catalytic kinase domain (Goodman et al., 2001, Oncogene 20:3969-3978). Syk is also overexpressed in anaplastie large cell lymphoma (ALCL), a very aggressive large T- or null-cell lymphoma that usually expresses anaplastie lymphoma kinase (ALK) (Kienle et al., 2005, J. Clin. Oncol. 23:3780-3792).
Ultraviolet radiation (UVR) from sunlight is a major etiologic factor in human skin photoaging and photocarcinogenesis. The ozone layer blocks UVC (180-280 nm). It is generally thought that UVB (280-320 nm) and UVA (320-400 nm) are responsible for sunlight-induced skin photodiseases (de Gruijl, 1999, Eur. J. Cancer 35:2003-2009; de Gruijl, 2000, Methods Enzymol. 319:359-366). UVA comprises approximately 90% or more of the total solar UVR, while UVB comprises the remaining 1-10% of the total UVR (de Gruijl, 1999, Eur. J. Cancer 35:2003-2009; Woodhead et al., 1999, J. Epidemiol. 9:S102-S114).
The signal cascade for UVR effects in skin begin with the stimulation of cell surface receptors, like epidermal growth factor receptor (EGFR). Following the activation of cell surface receptors by UVR, subsequent activation of mitogen-activated protein kinases (MAPKs) occurs by dual phosphorylation on threonine and tyrosine at Threonine-X-Tyrosine motifs within the activation loop of MAPKs. MAPKs are a family of proteins that include the extracellular signal regulated kinases (ERKs), p38 kinase, and c-Jun NH2-terminal kinase (JNKs) (Yousefi et al., 1996, J. Exp. Med. 183:1407-1414). ERK has two isoforms (1/2); p38 kinase has four isoforms (α/β/δ/γ); and JNKs has three isoforms (1/2/3) (Pearson et al., 2001, Endocr. Rev. 22:153-183; Tibbles et al., 1999, Cell. Mol. Life. Sci. 55:1230-1254).
One or more MAPK kinases (MAPKKs) catalyze this phosphorylation. MAPKKs themselves are activated by MAPKK kinases (MAPKKKs). Besides the cell surface receptor induced MAPKKK-MAPKK-MAPK pathway, MAPK can also be activated through the Ras family of G proteins. After Ras is activated, it will initially induce cell proliferation, targeting MEK1/2 MAPKK activation. Activated MAPKKs further activate ERK1/2 that can induce the AP-1 transcription factor that is related to cell proliferation. Ras-MEK-ERK also activates p38 and increases the expression of p53 and p16, which have anti-proliferation function (Iordanov et al., 2002, Mol. Cell. Biol. 22:5380-5394; Lin et al., 1998, Genes Dev. 12:3008-3019).
Normally, MAPKs mediate several cellular and organismal functions including proliferation, growth, differentiation, development, and apoptosis. MAPKs also play a significant role in mediating the UV induced biological effects, making the MAPK signaling cascade the key signal pathway triggered by UV (Bode et al., 2003, Sci. SETK 167:RE2; Kyriakis et al., 2001, Physiol. Rev. 81:807-869). Under stress conditions like UV stimulation, MAPK signaling is important for protecting the epidermis against UVR induced skin damage and carcinogenesis by activating cell cycle arrest, apoptosis, and inflammation in damaged tissues (Xia et al., 1995, Science 270:1326-1331; Chang et al., 2001, Nature 410:37-40).
Matrix metalloproteinases (MMPs) are thought to play an important role in the pathology of photoaging (Fisher et al., 1996, Nature 379:335-339; Scharffetter et al., 1991, Arch. Dermatol. Res. 283:506-511). Both UVA and UVB can induce MMP overexpression; however, several differences have been found between UVA and UVB induced signaling.
UVB is mostly absorbed by keratinocytes in the epidermis. After the transcription factors activator protein-1 (AP-1) and nuclear factor κB (NF-κB) are induced, they can further activate the gene expression of MMPs. In contrast to the UVA mediated signaling pathway, UVB may activate the MAPKs by stimulating the EGFR and/or PKC (a typical protein kinase C). UVB can also activate the PI3K (phosphatidylinositol 3-kinase) pathway, which can further activate many downstream molecules such as Akt (also called protein kinase B). In general terms, JNK and p38 MAPKs are activated by stress stimulations such as UVR while ERKs are activated by mitogenic stimuli (Xia et al., 1995, Science 270:1326-1331; Chang et al., 2001, Nature 410:37-40). However, there is significant cross talk among these three MAPKs p38 and ERKs).
In contrast, the longer wavelength UVA penetrates deeper into the skin and can affect both epidermal keratinocytes and dermal fibroblasts. UVA affects expression of MMPs mainly through the generation of reactive oxygen species (ROS) (Fisher et al., 1997, N. Engl. J. Med. 337; 1419-1428; Fisher et al., 1998, J. Clin. Invest. 101:1432-1440; Sato et al., 1993, Oncogene 8:395-405). It was first thought that UVA might phosphorylate EGFR, thus activating several downstream effector molecules, leading to the phosphorylation of p70S6K and p90RSK. These two phosphorylated proteins could further phosphorylate the 40S ribosomal protein S6. UVA might also activate the JNKs, ATM (ataxia telangiectasia imitated) and SMase (sphingomyelinase) which lead to apoptosis (Zhang et al., 2001, DNA Cell Biol. 20; 769-779; Zhang et al., 2001, J. Biol. Chem. 276:14572-14580; Hamilton et al., 1998, J. Biol. Chem. 273:28155-28162).
UVB was thought to be the highly mutagenic agent responsible for skin cancer development, because DNA is a chromophore for UVB radiation (but not for UVA) (Rosenstein et al., 1987, Photochem. Photobiol. 45:775-780). However, the UVA and UVB generated free radicals such as OH radical, one-electron oxidation oxidants and single oxygen can still damage DNA and cause base pair loss, single strand breaks, and protein-DNA crosslinking (Ichibashi et al., 2003, Toxicology 189; 21-39; Peus et al., 1998, J. Invest. Dermatol. 110:966-971; Darr et al., 1994, J. Invest. Dermatol. 102:671-675).
In the skin, many studies have demonstrated that MAPKs play an important role in regulating several key oncogenes and tumor suppressors that are relevant to UV induced cancers including p53, p16, APC/β-catenin, and Ha-Ras. JNK and p38 are thought to block cell proliferation or promote cell apoptosis via modulation of p53, which can prevent the tumor growing. Apoptosis would be impaired in the absence of JNK and p38 (Wang et al., 2002, Mol. Cell. Biol. 22:3389-3403; She et al., 2000, Oncogene 21:1580-1589; Hildesheim et al., 2004, J. Invest. Dermatol. 122:497-502). For instance, it was found that p38 inhibitor SB202190 blocked the UV induced apoptosis in mouse skin. p38 could phosphorylate Ser33 and ser37 of p53, while the Ser15 of p53 could be phosphorylated by p38 and ERK, and JNK phosphorylated Ser20 of p53 (She et al., 2000, Oncogene 21:1580-1589; Hildesheim et al., 2004, J. Invest. Dermatol. 122; 497-502; She et al., 2000, J. Biol. Chem. 275:20444-20449; Shieh et al., 2000, Genes Dev. 14:289-300).
MAPKs involved apoptosis is an essential and effective way to protect and repair UV induced skin damage. MAPKs are not only involved in apoptosis but also required by the UV induced inflammation. Keratinocytes are the major cells that release cytokines in the epidermis. After UV radiation, they secrete many cytokines, such as granulocyte colony-stimulating factor (G-CSF), IL-3, IL-8, macrophage-CSF, transforming growth factor α/β (TGF α/β), platelet-derived growth factor (PDGF), GM-CSF, interferon gamma (INFγ). These cytokines can further attract Langerhan's cells and activate neutrophils, macrophages and fibroblasts. MAPKs have been shown to regulate expression of all these cytokines (Ansel et al., 1990, J. Invest. Dermatol. 94:101 S-107S; Luger et al., 1990, J. Invest, Dermatol. 95:100 S-104S; Beyaert et al., 1996, EMBO J. 15:1914-1923; Lee et al., 1996, J. Leukoc. Biol. 59:152-157; Wery-Zennaro et al., 2000, Oncogene 19:1596-1604).
There remains a need in the art to identify novel markers of UVR damage in skin as well as new therapeutic targets for the treatment and remediation of UVR damage. The present invention fills this need.